RNA viruses are a significant cause of morbidity and mortality in humans. Some representatives of this large group of pathogens are hepatitis A, B and C viruses, HIV, influenza virus, most of the cold viruses, rabies virus, Ebola virus, West Nile virus, and dengue virus. We face major difficulties in controlling these viral diseases because of the viruses' escape from immune responses, escape from antiviral drugs, and the emergence of new viruses from animal reservoirs. All these difficulties are caused by the same process: RNA viruses rapidly adapt to new challenges in their environment. This rapid adaptation is possible because RNA virus populations are extremely heterogeneous. Among the variants in a viral population there are usually mutants capable of replicating in the presence of antibodies, antiviral drugs, or in new host cells. New therapies must be designed such that the development of resistance can be prevented or at least substantially delayed. However, we currently have no systematic strategy to develop such therapies, and the predominant approach is trial-and-error. This proposal will continue to build a framework of RNA virus evolution that will be used to develop improved antiviral strategies in a rational way. We use a unique approach of experimental virus evolution (which includes detailed phenotypic and genotypic analyses) combined with computer simulation to address fundamental questions of viral adaptation. We propose to study genetic and environmental determinants that lead to fitness changes and to differences in the potential of a virus population to adapt. Our model system is vesicular stomatitis virus, a well characterized and widely studied virus. We will evaluate three aspects of RNA virus evolution: (i) The genetic basis of differences in adaptability; (ii) the effect of two environmental determinants, population size and population density, on the speed of evolution and the maintenance of variation; and (iii) the consequences of interactions between genetic components and environmental components for virus evolution.